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IDENTIFICATION OF NOVEL ANDROGEN RECEPTOR
TARGET GENES IN PROSTATE CANCER
by
Unnati Jariwala
A Dissertation Presented to the
FACULTY OF THE GRADUATE SCHOOL
UNIVERSITY OF SOUTHERN CALIFORNIA
In Partial Fulfillment of the
Requirements for the Degree
DOCTOR OF PHILOSOPHY
(BIOCHEMISTRY AND MOLECULAR BIOLOGY)
August 2007
Copyright 2007 Unnati Jariwala

The androgen receptor (AR) has unequivocal roles throughout all phases of prostate cancer (PCa). The downstream target genes that mediate the receptor's oncogenic functions, however, remain ill-defined. This thesis describes studies undertaken to identify AR target genes in C4-2B advanced human PCa cells using two chromatin immunoprecipitation-based methodologies: ChIP Display and ChIP-on-chip.; Using ChIP Display we discovered 19 AR-occupied regions (ARORs). Our data suggests that AR does not prefer binding to 5'proximal-promoter sequences; on the other hand, it does tend to occupy intragenic sequences, mostly introns. Approximately half of the genes near ARORs were regulated by the AR, usually positively, and usually, but not always, in a ligand-dependent manner. Although the AR generally bound the same regions in the androgen-dependent LNCaP and the castrate resistant cell line C4-2B, we detected differences in how nearby genes are regulated. Finally, we provide evidence in support of AR-mediated regulation of some of the newly discovered AR target genes in vivo.; Using ChIP-on-chip we identified an additional 62 novel ARORs on chromosomes 19 and 20. Again, AR demonstrated no preference for binding to 5'gene-flanking sequences. Sixty percent of the 62 ARORs were intragenic. Most of the 62 ARORs were far from transcription start sites, overlapped with histone acetylation, and exhibited some conservation across species. Sixty percent of ARORs demonstrated enhancer activity in reporter assays. Several approaches we took to analyze gene expression showed that only 9/95 annotated genes in proximity to ARORs responded to DHT treatment. The use of high-resolution tiling microarray led us to identify TUF1 (transcript of unknown function 1), which was highly stimulated by DHT. Sequence analysis revealed that TUF1 encodes a putative collagenous protein with high degree of conservation.; Several genes we identified as AR targets (for example, PRKCD, PYCR1, JAG1 and MMP24) have been implicated in cancer progression or have functions which make them likely candidates for mediating AR's oncogenic functions. Future studies on the molecular mechanisms of these target genes in PCa will shed light on aspects of disease progression and may provide novel targets for drug development.

IDENTIFICATION OF NOVEL ANDROGEN RECEPTOR
TARGET GENES IN PROSTATE CANCER
by
Unnati Jariwala
A Dissertation Presented to the
FACULTY OF THE GRADUATE SCHOOL
UNIVERSITY OF SOUTHERN CALIFORNIA
In Partial Fulfillment of the
Requirements for the Degree
DOCTOR OF PHILOSOPHY
(BIOCHEMISTRY AND MOLECULAR BIOLOGY)
August 2007
Copyright 2007 Unnati Jariwala